YAP1通过结合TEAD2调控靶基因BIRC5参与人晶状体上皮细胞上皮-间质转化OA

Objective To explore the involvement of Yes-associated protein 1(YAP1)in the epithelial-mesenchymal transition(EMT)of human lens epithelial cells(LECs)by binding to transcriptional enhancer domain transcription factor 2(TEAD2)to regulate the target gene baculovirus inhibitor of apoptosis protein 5(BIRC5).Methods HLE-B3 cells were divided into the Vector group,OE-YAP1 group,and OE-TEAD2 group,and transfected with pCDNA3.1-Vector,pCDNA3.1-YAP1,and pCDNA3.1-TEAD2,respectively.The transfection was observed under an inverted fluorescence microscope.After successful transfection,the cultured HLE-B3 cells were further cultured for 24 hours using DEME with 10 μg·L-1 transfor-ming growth factor-β2 and divided into the Vector+T empty vector group,OE-YAP1+T transfection group,OE-TEAD2+T transfection group,and OE-YAP1+OE-TEAD2+T co-transfection group.Real-time fluorescence quantitative PCR and West-ern blot were used to detect the relative mRNA and protein expression levels of YAP1,TEAD2,BIRC5,E-cadherin,α-smooth muscle actin(α-SMA),and Vimentin in each group of cells.Wild-type vector pGL3-basic-BIRC5-WT containing binding sites and mutant vector pGL3-basic-BIRC5-MUT containing mutation sites were constructed and transfected with pCDNA3.1-TED2 or pCDNA3.1-Vector into HLE-B3 cells.The interaction between TEAD2 and BIRC5 was verified using the dual lucif-erase reporter gene assay.Results The overexpression plasmids YAP1 and TEAD2,as well as the empty vector,were highly expressed in cells in the Vector group,OE-YAP1 group,and OE-TEAD2 group,and can be used for subsequent experi-ments.The relative protein and mRNA expression levels of YAP1,TEAD2,and BIRC5 in the OE-YAP1+OE-TEAD2+T co-transfection group were higher than those in the Vector+T empty vector group and the OE-YAP1+T transfection group,and the differences were statistically significant(all P＜0.05).Compared with the Vector+T empty vector group and the OE-YAP1+T transfection group,the OE-YAP1+OE-TEAD2+T co-transfection group had lower protein and mRNA expression levels of E-cadherin and higher protein and mRNA expression levels of α-SMA and Vimentin,with statistically significant differences(all P＜0.05).The results of the dual luciferase reporter gene assay confirmed that TEAD2 could activate the transcription of BIRC5.Conclusion YAP1 can regulate the target gene BIRC5 to participate in the EMT of LECs by bind-ing to TEAD2,providing a new approach for the prevention and treatment of posterior capsular opacification.